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Asim et al. Journal of Biomedical Science (2015) 22:41 DOI 10.1186/s12929-015-0138-y

REVIEW Open Access “Down : an insight of the ” Ambreen Asim, Ashok Kumar, Srinivasan Muthuswamy, Shalu Jain and Sarita Agarwal*

Abstract Down syndrome (DS) is one of the commonest disorders with huge medical and social cost. DS is associated with number of phenotypes including congenital heart defects, , Alzeihmer’s disease, Hirschsprung disease etc. DS individuals are affected by these phenotypes to a variable extent thus understanding the cause of this variation is a key challenge. In the present review article, we emphasize an overview of DS, DS-associated phenotypes diagnosis and management of the disease. The or miRNA involved in Down syndrome associated Alzheimer’s disease, congenital heart defects (AVSD), leukemia including AMKL and ALL, hypertension and Hirschprung disease are discussed in this article. Moreover, we have also reviewed various prenatal diagnostic method from karyotyping to rapid molecular methods - MLPA, FISH, QF-PCR, PSQ, NGS and noninvasive prenatal diagnosis in detail.

Introduction content of SINE’s, LINE’s, and LTR are 10.84%, 15.15%, Down syndrome is one of the most leading causes of in- 9.21% respectively. The Table 1 given below highlights tellectual and millions of these patients face some of the genes present on 21. various health issues including learning and memory, congenital heart (CHD), Alzheimer’s diseases Features of DS (AD), leukemia, and Hirschprung disease(HD). There are various conserved features occurring in all DS The incidence of is influenced by maternal age population, including learning , craniofacial ab- and differs in population (between 1 in 319 and 1 in normality and in early infancy [13]. Some people 1000 live births) [1-5]. DS has high genetic complexity of DS are affected by variant phenotypes including atrio- and phenotype variability [6-8]. Trisomic fetuses are at ventricular septal defects (AVSD) in heart, leukemia’s(both elevated risk of and DS people have in- acute megakaryoblastic leukemia(AMKL) and acute creased incidence of developing several medical condi- lymphoblastic leukemia(ALL)), AD and HD. DS individual tions [9]. Recent advancement in medical treatment with have variety of physical characteristics like a small chin, social support has increased the life expectancy for DS slanted eye, poor muscle tone, a flat nasal bridge, a single population. In developed countries, the average life span crease of the palm and a protuding due to small mouth for DS population is 55 years [10]. and large tongue [14]. Other features includes big toe, abnormal pattern of fingerprint and short fingers. Review Human of the disease DS complex phenotype results from dosage imbalance of The most common cause of having a DS babies is pres- genes located on human chromosome 21(Hsa 21). The ence extra copy chromosome 21 resulting in trisomy. genetic nature of DS together with the relatively small The other causes can be Robertsonian translocation and size of Hsa 21 encouraged scientist to concentrate efforts isochromosomal or . Ischromosome is towards the complete characterization of this chromo- a term used to describe a condition in which two long some in the past few years. The length of 21q is 33.5 Mb arms of chromosome separate together rather than the [11] and 21 p is 5–15 Mb [12]. A total 225 genes was es- long and short arm separating together during egg timated when initial sequence of 21q was published [11]. sperm development. Trisomy 21 ( 47, XX, + 21 Hsa 21 has 40.06% repeat content out of which the repeat for females and 47, XY, + 21 for males) is caused by a failure of the chromosome 21 to separate during egg or * Correspondence: [email protected] Department of , Sanjay Gandhi Post Graduate Institute of sperm development. In Robertsonian translocation Medical Sciences, Lucknow 226014, India which occurs only in 2-4% of the cases, the long arm of

© 2015 Asim et al. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http:// creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Asim et al. Journal of Biomedical Science (2015) 22:41 Page 2 of 9

Table 1 Some common present in chromosome 21 18]. The sequencing of Hsa 21 proved to be an import- S. no. Genes Abbreviation ant factor in the progression of DS research [19] and led 1. APP (A4) precursor to further insight into -phenotype correlations 2. C21 or f59: Chromosome 21 open reading frame 59 associated with DS and precise characterizations of DSCR regions [13]. A “critical region” within 21q22 was 3. CBS Cystathionine-beta-synthase believed to be responsible for several DS phenotypes 4. CLDN14 Claudin 14 including craniofacial abnormalities, congenital heart de- 5. HLCS Holocarboxylase synthetase (biotin-(proprionyl- fects of the endocardial cushions, of the coenzyme a-carboxylase (ATP-hydrolysing)) ligase) fifth finger and mental retardation [20]. 6. KCNE1 Potassium voltage-gated channel, Dual-specificity tyrosine phosphorylation-regulated kin- isk-related family, member 1 ase (DYRK1A) and regulator of calcineurin 1 (RCAN1), 7. KCNE 2 Potassium voltage-gated channel, Down syndrome molecule (DSCAM) has isk-related family, member 2 been suggested to play a critical role in the developing 8. LAD Leukocyte adhesion deficiency brain and has also been identified as a candidate gene for 9. SOD 1 1 the increased risk of CHD in DS individuals [21,22]. 10. TMPRSS3 Transmembrane protease, serine 3 DSCAM is a critical factor in neural differentiation, axon 11. PCNT Centrosomal pericentrin guidance, and the establishment of neural networks and it 12. DSCR1 Down Syndrome critical region 1 has been suggested that the disruption of these processes contributes to the DS neurocognitive phenotype [22]. 13. DYRK1A Dual specificity tyrosine-(Y)-phosphorylation regulated kinase 1A Based on thorough analyses of studies on humans and DS 14. RRPB1 Ribosomal RNA processing 1 homolog B mouse models, it is evident that there is not a single crit- ical region of genes sufficient to cause all DS phenotypes. 15. Calcium binding protein Alternatively, it is likely that there are multiple critical re- gions or critical genes contributing to a respective pheno- type or group of phenotypes associated with DS [23]. the chromosome 21 is attached to another chromosome (generally ). While mosaicism deals with Various clinical conditions associated to Down syndrome the error or misdivision occurs after fertilization at some The various clinical conditions associated with DS are point during cell division. Due to this people with mo- Alzheimer’s disease, heart defects, leukemia, hyperten- saic DS have two cell lineages which contribute to tis- sion and gastrointestinal problems (Figure 1). The mo- sues and organs of individuals with Mosacism (one with lecular pathogenesis mechanism of these DS related the normal number of , and other one phenotype must be studied along with its causative with an extra number 21) [15]. agents in order to have a better understanding of the dis- ease. Below are some DS related phenotype discussed in Genotype-phenotype correlation detail which are as follows: Gene dosage imbalance hypothesis states that DS pa- tients have an increased dosage or copy number of genes Neurological problems on Hsa 21 that may lead to an increase in gene expres- DS patients have greatly increased risk of early onset AD. sion [13-15]. This hypothesis has been extended to in- After the age of 50, the risk of developing dementia in- clude the possibility that specific genes or subsets of creases in DS patients up to 70% [23-27]. There are vari- genes may control specific DS phenotypes [16]. Ampli- ous genes reported to cause early onset AD. Some of the fied developmental instability hypothesis states that a genes described in the current literature are APP (amyloid non-specific dosage of a number of trisomic genes leads precursor protein), BACE2 (beta secretase 2), PICALM to a genetic imbalance that causes a great impact on the (Phosphatidylinositol binding clathrin assembly protein) expression and regulation of many genes throughout the and APOE(Apolipoprotein E) etc. APP is an integral [13, 14]. Another hypothesis known as critical membrane protein which is concentrated in synapse of region hypothesis was also added to this list. Phenotypic and trisomy of this protein is likely to make sig- analyses was done on individuals with partial trisomy for nificant contribution to the increased frequency of demen- Hsa21 identified that only one or a few small chromo- tia in DS individuals. The triplication of Hsa 21 along with somal regions, termed “Down syndrome critical regions” APP in people without DS has been recently shown to be (DSCR) a region of 3.8-6.5 Mb on 21q21.22, with ap- associated with early onset AD. A tetranucleotide repeat, proximately 30 genes responsible for the majority of DS ATTT , in intron 7 of the amyloid precursor protein has phenotypes [15,16]. Previously a region of 1.6 to 2.5 Mb been associated with the age of onset of AD in DS in a was recognised as sufficient cause for DS pehnotype [17, preliminary study [28]. Various mouse models are used to Asim et al. Journal of Biomedical Science (2015) 22:41 Page 3 of 9

observe degeneration of basal forebrain cholinergic neu- with complete AVSD suggests the defects in CRELD 1 rons (BFCNs). Ts65Dn mice is dependent on trisomy of contribute to pathogenesis of AVSD in context with APP expression of retrograde axonal transport [29]. Stud- trisomy 21. ies have also revealed that BACE2 which encodes enzyme beta secretase 2 is also involved in AD. APP and BACE 2 Hematological problems genes are located on chromosome 21. The current data on Patients with DS display a unique spectrum of malignan- DS support the association of haplotypes in BACE2 with cies, which include leukemia’saswellassolidtumors.The AD [30]. Besides APP and BACE2 genes, other genes like first report of leukemia in a DS patient occurred in 1930 PICALM and APOE are also found to be associated with [39] and the first systematic study in 1957 [40]. Studies the age of onset of Alzheimer’s dimentia in DS [31]. indicate that patients with DS have a 10–20 fold in- creased relative risk of leukemia, with a cumulative risk Cardiac problems of2%byage5and2.7%byage30[41].Theyconstitute The incidence of CHD in newborn babies with DS is up approximately 2% of all pediatric acute lymphoblastic leu- to 50% [32]. Endocardial cushion defect also called as kemia(ALL) and approximately 10% of pediatric acute atrioventricular cushion defect is most common form myeloid leukemia (AML). Leukemogenesis of acute mega- which affects up to 40% of the patients. Ventricular sep- karyoblastic leukemia (AMKL) in DS patients is associated tal defect (VSD) is also present in these population with the presence of somatic involving GATA 1 which affects up to 35% of the patients [33]. The essen- gene (or also called as GATA-binding factor 1) [42]. tial morphological hallmark of an AVSD is the presence GATA 1 is a chromosome X- linked transcription factor of a common atrioventricular junction as compared to which is essential for erythoid and megakaryocytic differ- the separate right and left atrioventricular junction in entiation. Because of these GATA 1 mutations, there is a the normal heart. Other morphological features include production of shorter GATA 1 protein thereby leading to defects of the muscular and membranous atrioventricu- uncontrolled proliferation of immature lar septum and an ovoid shape of the common atrioven- [42,43]. On the other hand, acquired gain of function mu- tricular junction. There is disproportion of outlet and tation in Janus Kinase 2 gene are present in approximately inlet dimensions of the left ventricle, with the former 30% of cases with ALL in DS [44,45]. greater than the latter as compared to the normal heart where both dimensions are similar [34]. While in case of Hypertension VSD, the defect lies in ventricular septum of the heart People with DS have been reported to have a reduced inci- due to which some of the blood from the left ventricle dence of hypertension [46,47]. Trisomy of the Hsa21 leaks into the right ventric leading to pulmonary hyper- microRNA hsa-miR-155 contributes to this [48]. Hsa- tension. in non Hsa 21 CRELD1 (Cysteine rich miR-155 is proposed to specifically target one allele of the EGF like domain1) gene contributes to the development type-1 angiotensin II receptor (AGTR1) gene, resulting in of AVSD in DS [35]. CRELD1 is located on chromosome it’s under- expression, which contribute to a reduced risk 3p25. It encodes a cell surface protein that functions as of hypertension. Further studies are required to validate cell adhesion molecule and is expressed during cardiac this hypothesis and determine whether other genes may cushion development. CRELD1 gene contains 11 exons also protect people with DS against hypertension. spanning approximately 12 kb [36]. To the present, two specific genetic loci for AVSD have been identified. One Gastrointestinal problems was AVSD 1 locus present on chromosome 1p31-p21 DS patients constitute ~12% of all cases of HD. Duodenal [37]. Other locus was present on chromosome 3p25 and stenosis (DST) and (IA) are 260 and 33 the corresponding gene was CRELD1 [36,38]. Maslen et times more likely to occur DS [23,49]. HD is a form of low al. in [33] have identified two heterozygous missense intestinal obstruction caused by the absence of normal mutation (p.R329C and p.E414K) with two subjects in myenteric ganglion cells in a segment of the colon [50]. In DS and AVSD. They have recruited 39 individual of DS HD children, the absence of ganglion cells results in the with complete AVSD and have found the same mutations. failure of the distal intestine to relax normally. Peristaltic In the same study, DNA of 30 individual of trisomy without waves do not pass through the aganglionic segment and CHD was studied for both mutations, no such mutation there is no normal defecation, leading to functional was identified [35]. R329C which was originally reported in obstruction. Abdominal distention, failure to pass meco- an individual with sporadic partial AVSD and now it is also nium, enterocolitis and bilious vomiting are the predomin- detected in individual of DS with AVSD. Interestingly, with ant and appear within a few days thesamemutation(p.R329C),the severity of heart defect after birth. Infants with or DST present was greater in patients of DS with AVSD. Thus, identifica- with bilious vomiting early in the neonatal period. If left tion of CRELD 1 mutation in 2/39 individual (5.1%) of DS untreated, it will result in severe dehydration and Asim et al. Journal of Biomedical Science (2015) 22:41 Page 4 of 9

electrolyte imbalance. IA is a birth defects in which the rec- Simultaneously parental origin of the nondysjunction can tum is malformed and it is associated with an increased in- also be detected [57,58]. Additional method to measure cidence of some other specific anomalies as well, together copy number of DNA sequences include MLPA [59] which being called the VACTERL association: vertebral anomalies, was first introduced in 2002 as a method of relative quanti- anal atresia, cardiovascular anomalies, tracheoesophageal fication in DNA. MLPA offers various advantages like – a fistula, esophageal atresia, renal and limb defects. very short time for diagnosis (2–4 days), effectiveness, sim- Alterations of approximately 10 non Hsa21 genes have plicity and relatively low costs. It is based on hybridization been linked to this disease [51]. Several researches have and PCR method and is divided into four steps: DNA de- shown that HD contain the DSCAM gene which is naturation, hybridization of probe to the complementary expressed in neural crest that give rise to enteric ner- target sequence, probe ligation and PCR amplification. And vous system [49]. Overlapping critical region was de- finally capillary electrophoresis of PCR amplified products scribed both for DST and IA [51]. No other Hsa21 genes is carried out. However MLPA is unable to exclude low have been implicated so far. level placental and true mosaicism [60].

Diagnostic methods Advancement in the diagnosis Prevention of DS depends upon offering prenatal diagno- A recent method, termed paralogous sequence quantifica- sis to high risk via and chori- tion (PSQ), uses paralogous sequences to quantify the Hsa onic villus sampling (CVS). Amniocentesis and CVS are 21 copy number. PSQ is a PCR based method for the de- quite reliable but offers risk of of between 0.5 tection of targeted chromosome number abnormalities to 1% [52]. Based soft markers like small or no nasal bone, termed paralogous sequence quantification (PSQ), based large ventricles and nuchal fold thickness, the risk of DS on the use of paralogous genes. Paralogous sequences have for fetus can be identified through ultrasound generally at a high degree of sequence identity, but accumulate nucleo- 14 to 24 weeks of gestation [53]. Increased fetal nuchal tide substitutions in a locus specific manner. These se- translucency indicates an increased risk of DS [54]. The quence differences, which are termed as paralogous other methods used for prenatal diagnosis in which sequence mismatches (PSMs), can be quantified using py- traditional cytogenic analysis is still widely used in diffe- rosequencing technology, to estimate the relative dosage rent countries. However some rapid molecular assays- between different chromosomes. PSQ is a robust, easy to FISH(fluorescent in situ hybridization), QF-PCR (quantitative interpret, and easy to set up method for the diagnosis of fluorescence PCR), and MLPA(multiplex probe ligation common , and can be performed in less than assay)- also used for prenatal diagnosis. 48 h, representing a competitive alternative for widespread use in diagnostic laboratories. The sequencing is quantita- Routine karyotyping tively done by using pyrosequencing [61]. Finally, com- Cytogenetic analysis of metaphase karyotype remains the parative genomic hybridization (CGH) on BAC chips can standard practice to identify not only trisomy 21, but be used for the diagnosis of full trisomy or , also all other aneuploidies and balanced translocations. and for partial (segmental) aneuploidies [62,63]. Details on diagnostic methods with advantages and dis- advantages are mentioned in Table 2. Noninvasive Prenatal diagnosis Fetal cells in maternal ciruculation: Ever since the dis- Rapid testing methods covery of presence of fetal in maternal Over the past 10 years however, several other methods blood was made in 1969, the investigators are trying to have been developed and used for the rapid detection of develop genetics-based noninvasive prenatal diagnostics trisomy 21, either in fetal life or after birth. The most (NIPD) [64]. Despite several advantages offered by this widely used is FISH of interphase nuclei, using Hsa 21- approach, the use of fetal cells for NIPD has never specific probes or whole-Hsa 21 [55]. An alternative reached clinical implementation because of their paucity method that is now widely used in some countries is QF- (on the order of a few cells per milliliter of maternal PCR, in which DNA polymorphic markers (microsatellites) blood) and concerns of fetal cell persistence in the ma- on Hsa 21 are used to determine the presence of three dif- ternal circulation between pregnancies. ferent alleles [56]. This method relies on informative Cell free fetal DNA from maternal serum: This novel markers and the availability of DNA. Rapid diagnosis by approach was proposed in 1997. Cell-free fetal DNA PCR-based methods using polymorphic STR markers may constitutes between 5% and 10% of the total DNA in reduce these difficulties using conventional approach. Using maternal plasma and increases during gestation and rap- STR markers method we can detect trisomy in 86.67% idly clears from the circulation post delivery. Several cases with only two markers. Using more number of clinical applications based on the analysis of cell-free markers can further increase the reliability of the test. fetal DNA have been developed like determining fetal Asim et al. Journal of Biomedical Science (2015) 22:41 Page 5 of 9

Table 2 Common techniques used for diagnosis of Down’s syndrome along with its advantages and disadvantages Method Description Advantages Disadvantages 1 analysis Giemsa banding (G-banding) is • Suitable for low income • Time consuming. performed on fetal cells at countries where physician • Resolution of special importance for metaphase stage on amniocytes can be presumed to have (grown in vitro) or CVS. acquired a high level of diagnostic the detection of structural. skill in the absence of abnormalities may be quite low as the spontaneous dividing cells are laboratory services. more condensed than those obtained after cell culture in vitro. • In CVS, occurrence of confined placental mosaicism and occurrence of aberrant cells that do not represent the status of fetus. • Chances of giving a false positive and false negative result. 2 FISH(Fluorescence in FISH involves hybridization of • As it uses smaller probes thus • Sometimes diffused signals are situ hybridization) selected chromosome specific the signals appears to be more obtained because it uses chromosome DNA sequences that have been distinct as dots. at interphase stage which appears less labeled with fluorescent dye to condense than those of metaphase. chromosome preparation. The • • fluorescently labeled sequences It uses higher number of Time consuming since it involves stick to corresponding DNA of interphase nuclei for analysis, preparation of slides, fluorescent so the problem of any suspected microscopy and spot counting chromosome and can be visualized mosaicism is resolved. (~30min per sample is expected). under microscope. • Maternal and fetal XX is not distinguished by FISH. 3 QF-PCR (Quantitative Involves amplification and detection • Highly reliable and reproducible. • Poses a challenge in the case of fluorescent-polymerase of STR using fluorescently labeled mosaicism. chain reaction) primers. The product is thus visualized • Chances of getting false negative • While testing sex chromosome and quantified as peaks areas of and false positive cases are rare. abnormalities samples from normal XX respective length using an automated female may show homozygous DNA sequencer with Gene Scan • Maternal contamination is software. QF-PCR pattern indistinguishable from easily detected. those produced by sample with single • Faster approach as it can give XasinTurnersyndrome. the diagnosis within 24 hours. 4 Paralogous sequence A PCR based method for detection • The first generation design of • Expensive when compared to others. quantification (PSQ) of targeted chromosome number test requires 10 separate PCR abnormalities, based on the use reaction per sample, which of paralogous genes. Paralogous significantly reduces the sample sequences have high degree of throughput and increases the sequence identity but accumulate probability of handling errors. nucleotide substitution in a locus • It can handle 30–40 samples specific manner. These differences are called as paralogous sequence in a day and report result in mismatches which can be quantified less than 48 hours. using pyrosequencing. 5. MLPA (multiplex probe MLPA is based on hybridization and • Very short time for diagnosis • Unable to exclude low level ligation assay) PCR method. Divided into 4 phases: (2–4 days). placental and true mosaicism. DNA denaturation, hybridization of probe • to the complementary target sequence, Relatively low costs probe ligation and PCR amplification of ligated probe. These amplified products are analysed through capillary electropheresis. 6. NGS (Next Generation Clonally amplified DNA templates • The current time for sample • The cost of sequencing is approximately Sequencing) are sequenced in a massively parallel. processing, sequencing, and $700 –$1000 per sample. It provides a digital quantitative data interpretation in experienced • information, in that each sequence hands is 5 to 8 days. Complex data analysis. read is a countable “sequence tag” representing an individual clonal DNA template or a single DNA molecule. Asim et al. Journal of Biomedical Science (2015) 22:41 Page 6 of 9

Rh D status in Rh D-negative women [65], sex in sex- of clonally amplified DNA templates (or, most recently, linked disorders [66,67], and detection of paternally single DNA molecules) are sequenced in a massively inherited autosomal recessive and dominant mutations parallel fashion within a flow cell [72,73]. NGS provides [68]. However, there remains the outstanding challenge digital quantitative information, in which each sequence of the use of cell-free fetal DNA for the detection of read is a countable “sequence tag” representing an indi- chromosomal aneuploidy, in particular 21, 18, vidual clonal DNA template or a single DNA molecule. and 13. Several approaches have been adopted like the This quantification allows NGS to expand the digital origin of circulating cell-free fetal DNA is primarily the PCR concept of counting cell-free DNA molecules. placenta, whereas maternal cell-free DNA is derived from Fan et al. and Chiu et al. in 2008 described noninva- maternal leukocytes [69]. The approach includes studying sive detection of trisomy 21 by NGS [74]. Both groups differences in genomic DNA methylation between the pla- extracted cell-free DNA from maternal plasma samples centa and paired maternal leukocytes, investigators have from both euploid and trisomy pregnancies. DNA from characterized placenta-specific epigenetic markers [70] and each sample was sequenced on the Illumina Genome also finding of circulating cell-free placenta-derived mRNA Analyzer, and each sequence read was aligned to the ref- allowed the identification of placenta-specific mRNA pro- erence . Chiu et al. build on their earlier duction [71]. work with the Illumina Genome Analyzer and demonstrate The concept of digital PCR was also introduced to serve noninvasive NGS-based trisomy 21 detection with the the same purpose. In digital PCR, individual fetal and ma- sequencing-by-ligation approach on the Life Technologies ternal circulating cell-free DNA fragments are amplified SOLiD platform [75]. Cell-free DNA was extracted from 15 under limiting-dilution conditions and the total number of pregnant women, 5 of whom carried trisomy 21 fetuses and chromosome 21 amplifications (representing maternal plus it was clonally amplified by emulsion PCR, and sequenced fetal contributions) divided by the number of reference in 1 chamber of an 8-chamber SOLiD slide. This process chromosome amplifications should yield a ratio indicating yielded a median of 59 × 106 50-base reads per sample. A an over- or underrepresentation of chromosome 21. median of 12 × 106 reads (or 21%) were each aligned Although the digital PCR approach is conceptually uniquely to one location of the reference human genome solid, the low percentage of cell-free fetal DNA in the (with masking of repeat regions), for a coverage of approxi- maternal plasma sample requires the performance of mately 20% of the haploid human genome. For each tri- thousands of PCRs to generate a ratio with statistical somy 21 case, the chromosome 21 z score value indicated a confidence. This can be overcome by using of multiple 99% chance of a statistically significant difference from the target amplifications and enrichment of cell-free fetal chromosome 21 z scores for the controls. As reported earl- DNA which are still under research trail. ier with the Illumina Genome Analyzer, a nonuniform dis- Next recent method added to the list is next gener- tribution of aligned sequence reads was observed with the ation sequencing (NGS) which is based on the principle SOLiD data.

Figure 1 Various conditions associated with Downs’s syndrome with its causative genes. Asim et al. Journal of Biomedical Science (2015) 22:41 Page 7 of 9

The current time for sample processing, sequencing, Conclusion and data interpretation in experienced hands is 5 to 8 DS or Trisomy 21, being the most common chromosomal days for the Genome Analyzer and SOLiD platforms re- abnormality among live born infants, is associated with a spectively with the cost of approximately $700 – $1000 number of congenital malformations. Several theories have per sample. Going forward, one can expect streamlining been put forward to increase our understanding in pheno- and automation of technical processes and data analysis, type and genotype correlation. A “critical region” within coupled with reduced sequencing costs. 21q22 was believed to be responsible for several DS pheno- Ultimately, reduced sequencing costs and turnaround types including craniofacial abnormalities, congenital heart times could pave the way for NGS-based NIPD to be con- defects of the endocardial cushions, clinodactyly of the fifth sidered as an alternative to serum biomarker , finger and mental retardation and several other features. which,while cost-effective remains prone to false positives. The primary goal of this review is to unravel the common Forty years after the discovery of circulating fetal cells, the genes involved in DS associated phenotypes, including vision of NIPD appears clearer and closer. APP, BACE2, PICALM, APOE, GATA 1, JAK 2, CRELD 1 and DSCAM. This reviews also provides the detailed de- Management of the disease scription on the application of techniques to prenatal diag- One of the hallmarks of DS is the variability in the way nosis in DS. Rapid aneuploidy testing has been introduced that the condition affects people with DS. With the third in mid 1990’s in the form of FISH where testing can be 21st chromosome existing in every cell, it is not surprising done on uncultured amniocytes. Within a couple of years, to find that every system in the body is affected in some MLPA and QF-PCR has been added in the list of rapid an- way. However, not every child with DS has the same prob- euploidy testing. The other methods includes: NGS for cell lems or associated conditions. Parents of children with DS free fetal DNA screening for maternal plasma. Except ,FISH, should be aware of these possible conditions so they can MLPA and QF-PCR other method are not commercialized be diagnosed and treated quickly and appropriately. The for aneuploidy diagnosis due to their running cost, labor in- goal of the study is to point out the most common prob- tensive protocol and complex data analysis. Since various lems of which parents should be aware. clinical conditions are associated with DS, hence the man- Timely surgical treatment of cardiac defects during agement of these patients requires an organized multidiscip- first 6 months of life may prevent from serious compli- linary approach and continuous monitoring of these cations. Congenital occur in about 3% of chil- patients which has been discussed in this review article. dren and must be extracted soon after birth to allow light to reach the retina. A balance diet and regular exer- Competing interests cise are needed to maintain appropriate weight. Feeding The authors declare that they have no competing interests. problems and failure to thrive usually improve after car- Authors' contribution diac surgery. 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